OVERALL HYPOTHESIS: Signaling efficiency/specificity for heterotrimeric G-protein systems is determined in part by accessory proteins which regulate the efficiency and/or specificity of signal transfer from G-protein coupled receptors ^GPCRs) to G-proteins, segregate a signaling complex to microdomains of the cell, regulate the basal activity of the system and/or provide alternative modes of signal input to G-protein signaling systems that operate independent of a typical GPCR. Our objectives are to define such accessory proteins, their mechanism of regulation, their dysfunction in various diseases and their potential as therapeutic targets. We recently identified a group of proteins (Activators of G-protein signaling (AGS) 1-8 that directly influence the activation state of G-proteins independent of a GPCR. AGS proteins interact with different subunits and/or conformations of heterotrimeric G-proteins and selectively regulate different types of G-proteins. This proposal focuses on AGS1, which is unique among the AGS family members. AGS1 is a Ras related protein that regulates tieterotrimeric G-protein signaling providing a surprising mechanism for cross talk between small and large (heterotrimeric) G-protein families. We have little understanding of how AGS1 integrates into cell and organ function and a major goal of this application is to address this issue. SPECIFIC AIM #1 Define the subcellular location and regulation of AGS1. SPECIFIC AIM #2 Identify and characterize downstream signaling mechanisms of AGS1. SPECIFIC AIM #3 Determine the functional role of AGS1 in cell differentiation. AGS1 and related accessory proteins provide unexpected mechanisms for regulation of the G-protein activation cycle and have opened up a new area of research related to the cellular role of G-proteins as signal transducers and the cellular functions they regulate. As such, these proteins and the concepts advanced with their discovery provide unexpected avenue; for therapeutic development and increased understanding of disease mechanisms. [unreadable] [unreadable]